Lead is a ubiquitous environmental contaminant which causes neurological disorders at low exposure levels. Determination of the mechanisms of lead transport across the plasma membrane may be a first step in the development of new strategies to prevent toxicity in exposed individuals and redistribution of lead to soft tissues during chelation therapy. The specific aims of this proposal are: 1) to determine the contribution of lead entry via the cation channels involved in capacitative Ca2+ influx, 2) to test the involvement of a plasma membrane Ca2+ ATPase (PMCA) in lead efflux, and 3) to localize lead in live cells. Lead uptake into GH3, C6, and 301 cell lines and primary cultures of bovine brain endothelial cells will be measured using indo-1, an intracellular fluorescent dye which quenches upon binding lead Fluorescence will be measured with a fluorimeter, with single cell digital fluorescence imaging, and with confrontal microscopy. Lead uptake will be monitored in the presence of activators and inhibitors of nonvoltage sensitive Ca2+ channels. Transport through other cation channels will also be assessed by monitoring lead uptake in the presence of various cations. Lead efflux will be studied in the presence of activators and inhibitors of PMCA. Preliminary data indicate lead enters cells via a cation channel which is activated by the emptying of intracellular Ca2+ pools.